Accumulation of A plaque in the brain is a hallmark feature of Alzheimer's disease and a biomarker of disease progression. Observations in humans show that plaques are found in regions of the brain that display high levels of neuronal activity, sometimes referred to as the default mode network. Similarly, about 10% of individuals with temporal lobe epilepsy develop plaques within affected areas as early as 30 years of age. Studies from our lab have demonstrated that direct modulation of synaptic activity dynamically regulates brain A levels in awake animals, with increased synaptic activity rapidly increases brain interstitial fluid (ISF) A levels and vice versa for suppressed activity. These findings strongly suggest a close temporal relationship between synaptic activity and A generation. Determining the mechanisms that underlie this link remains important for understanding the pathological development of AD. We have developed novel micro- immunoelectrode electrodes (MIEs) that detect A with very high temporal resolution (measures A in vivo every minute). This approach enables us to study the rapid kinetics and dynamics of A in vivo. In our published studies (Prabhulkar et al. 2012), and in preliminary data we show that these MIEs can specifically measure ISF A1-40, A1-42 or aggregates, depending on the antibody attached to the electrode surface. Our in vivo MIE studies demonstrate brain interstitial fluid (ISF) A levels change from minute-to-minute in APP/PS1 transgenic mice. Previous publications from our group and others demonstrate that ISF A levels are closely linked to synaptic transmission. In vitro data suggest that high concentration and low pH facilitate conversion of A into toxic aggregates. Synaptic activity increases A generation within endosomes, a confined location where the pH is low, and the concentration of A can potentially be elevated. We propose that there is a rapid link between synaptic transmission and A generation, with higher frequencies of synaptic transmission causing more A to be formed. In addition, elevated synaptic A generation in low-pH endosomes will convert A into aggregated species (either oligomers or fibrils). The goal of this proposal is to use this new MIE technology in combination with pharmacological manipulation to block or enhance specific aspects of synaptic activity to elucidate the cellular mechanisms that regulate A generation and aggregation on a short time-scale. The results of these studies will improve our understanding of the temporal relationship between synaptic activity and A generation/species and uncover mechanisms involved in the progression of AD.